Among the apoptotic pathways involving p53-independent tumor necrosis factor receptors (TNFRs), cell death through the death receptor 5 (DR5, TRAIL receptor 2, TRICK2) or the death receptor 4 (DR4, TRAIL receptor 1) pathway activated by TNF-related apoptosis inducing ligand (TRAIL) has been a very attractive target for cancer therapy as they can specifically induce cancer cell death with little adverse side effects on normal cells (Ashkenazi et al., J. Clin. Invest., 104:155-162, 1999; and Ashkenazi, Nat. Rev. Cancer, 2:420-430, 2002).
Currently, there exist several methods to develop caner cell-specific therapeutic agents which target DR4 or DR5 such as a method using a recombinant TRAIL (for example, 114th to 281st amino acid residues of TRAIL) as a ligand of the death receptor and a method of developing an agonistic antibody among mouse- or human-derived complete antibodies (e.g.: mAb or IgG) specific to the death receptor (Pollack et al., Clin. Cancer Res., 7:1362-1369, 2001; Jo et al., Nat. Med., 6:564-567, 2000; Ichikawa et al., Nat. Med., 7:954-960, 2001; and Walczak et al., Nat. Med., 5:157-161, 1999). However, the recombinant TRAIL is very unstable and tends to form a soluble oligomer, whose apoptotic activity is about 20 to 100 times lower, and it causes side effects such as cytotoxicity and immune reaction on normal cells such as astrocytes, hepatocytes and keratinocytes (Jo et al., Nat. Med., 6:564-567, 2000). Further, TRAIL is not capable of inducing more than 50% cell death of malignant tumor cells (Zhang et al., Cancer Gene Ther., 12:228-237, 2005). Hence, a cancer cell that can be killed by TRAIL is designated a TRAIL-sensitive cancer cell, and a cancer cell not killed by TRAIL, a TRAIL-resistant cancer cell.
There have been developed antibodies, which have the specific binding affinity to DR5 to induce the cell death: humanized antibodies developed from mouse-derived monoclonal antibodies such as TRA-8 (mouse IgG) (Walczak et al, Nat. Med., 5:157-161) and AD5-10 (mouse IgG) (Guo et al., J. Biol. Chem., 280:41940-41952, 2005), as well as human-derived monoclonal antibodies such as HGS-ETR2 (human IgG1) (Georgakis et al., Br. J. Haematol., 130:501-510, 2005) and KMTR2 (human IgG4) (Motoki et al., Clin. Cancer Res., 11:3126-3135, 2005).
However, the above antibodies induce apoptotic cell death of TRAIL-sensitive cancer cells but not TRAIL-resistant cancer cells. Further, the antibodies in the form of an antigen-binding fragment (Fab) or a single chain variable fragment (scFv) having a monovalent antigen binding site does not induce cell death of cancer cells (e.g.: KMTR2), while antibodies in the form of IgG having divalent antigen binding site (e.g.: HGS-ETR2 and AD5-10) show cytotoxicity or induced cell death when an IgG is used as a cross-linker (Chuntharapai et al., J. Immunol., 166:4891-4898, 2001; Motoki et al., Clin. Cancer Res., 11:3126-3135, 2005; and Wajant et al., Oncogene, 20:4101-4106, 2001). But, it has not been reported that an anti-DR5 antibody in the form of a scFv and a Fab induces cancer cell death.
Currently, there remains a question as to whether or not autophagy is the mechanism responsible for inducing cancer cell death (Kondo et al., Nat. Rev. Cancer, 5:726-734, 2005), and it has been reported that only specific compounds can kill cancer cells by an autophagic cell death pathway (Yu et al., Science, 304:1500-1502, 2004). However, there have been no reports of anti-DR5 mAbs which induce autophagic cell death.